Ultrasound-assisted preparation of protein–polyphenol conjugates and their structural and functional characteristics

Herein, ultrasound-assisted conventional covalent binding methods (alkali treatment, free radical mediation, and an enzymatic method) were used to prepare soybean protein isolate (SPI)-(-)-epigallocatechin gallate (EGCG) conjugates to investigate the enhancement effect of the ultrasound synergistic treatment. In addition, the influence of EGCG grafting on the structure and properties of SPI was evaluated via reactive group analysis, spectral analysis, surface hydrophobicity measurements, emulsification property assessment, and α-glucosidase inhibition analysis. The obtained results revealed that the enzymatic method produced the highest polyphenol grafting content among the conventional techniques. Meanwhile, ultrasound treatment increased the amount of grafted polyphenol species during the alkali treatment and free radical mediation procedure, decreased the grafting efficiency in the enzymatic method, and maximized the grafting efficiency during the alkali treatment. In addition, reactive group and spectral analyses demonstrated that EGCG formed C-N and C-S bonds with SPI and decreased the α-helix content in the protein structure, thereby increasing the molecular flexibility of SPI. It also produced hydrogen bonds and hydrophobic interactions, as demonstrated by the results of molecular docking. Furthermore, the EGCG grafting of SPI conducted under the ultrasound-assisted conditions endowed SPI with unique functional characteristics, including good emulsification and antioxidant properties and high α-glucosidase inhibitory activity, while the ultrasound-assisted alkali treatment resulted in the optimal functional properties. The results of this study provide new insights into the effective preparation of SPI-EGCG complexes with multiple functionalities, thereby expanding the scope of high-value SPI utilization.